INTRODUCTION:

Ethoxy Analogue of Curcumin (EAC) shows great promise in oncology, particularly in treating breast and oral cancers, where it has demonstrated five times more apoptotic activity than curcumin^1,2. However, its clinical use is limited by poor bioavailability due to low solubility, rapid metabolism, and degradation in the GI tract³. To overcome these barriers, buccal films have been explored as an alternative delivery method. These films dissolve in saliva and release EAC directly into the oral cavity, enhancing local cancer treatment while bypassing the GI tract. This study investigates the impact of polymer selection and concentration on EAC release from buccal films. Hydrophilic polymers, critical for film adhesion and drug release, can promote nearly complete EAC release within 30 minutes⁴.

The research aims to identify optimal polymer formulations to improve EAC bioavailability and therapeutic effects, particularly for oral cancers. By targeting the cancer site directly and minimizing systemic exposure, buccal films offer a promising strategy to enhance EAC's effectiveness while reducing side effects.

EXPERIMENTAL METHODS:

Materials:

EAC powder, obtained from SUCP after ethylation of pure curcumin, was used alongside hydrophilic polymers—Polyethylene oxide (PEO), Hyaluronic acid (HA), and Pectin—to prepare buccal films using the solvent casting method^5-7. Pectin, a natural polysaccharide, was utilized in its low-molecular-weight form (LMWP) with a degree of esterification below 50% and molecular weight under 50,000 Daltons, offering improved solubility and faster gel formation. Other materials included PEG 4000 as a plasticizer, arrowroot starch, methanol, glycerine, tartaric acid, mannitol, Poloxamer 907, isotonic phosphate buffer (pH 6.8), bovine mucosa membrane, and an agar plate.

Instruments:

The equipment used includes an FT-IR spectrometer, USP-II dissolution paddle apparatus, UV spectrophotometer, micropipette, vernier calipers, electronic weighing balance, incubator, mortar and pestle, petri dish, 0.71 mm sieve, forceps, scissors, ruler, magnetic stirrer, rubber band, volumetric flasks (50 mL and 100 mL), heat plate with stirrer, measuring cylinder (10 mL), and beakers (100 mL, 200 mL, and 1 L).

Preparation of EAC Primary Standard Solutions:

To prepare the EAC Standard Stock Solution, 10 mg of EAC is dissolved in a 100 mL volumetric flask with isotonic phosphate buffer (pH 6.8). Next, 1 mg of cetrimide is added and mixed, then the solution is adjusted to 100 mL, resulting in a 100 µg/mL concentration.

Preparation of EAC Working Standard Solutions:

The working standard solution was prepared by diluting the primary stock solution with isotonic phosphate buffer (pH 6.8) in 10 mL volumetric flasks. Aliquots of 1-10 mL of the stock solution were pipetted into separate flasks and diluted to final concentrations ranging from 10 to 100 µg/mL.

Construction of Standard Calibration Curve for EAC-Dilutions:

A standard calibration curve for EAC dilutions was constructed by measuring absorbance at 425 nm using a UV-Vis spectrophotometer. Absorbance (y-axis) was plotted against concentration (x-axis), with linear regression confirming linearity (R² > 0.99). This curve is essential for determining EAC concentrations in unknown samples for solubility and release studies. (Figure-1)

Figure 1: Absorbance at 421 nm against concentration of curcumin solution (µg/mL)

Spectral Analysis of Stock solutions:

Absorbance was measured at 421 nm with a UV spectrophotometer against a buffer blank, creating a standard calibration curve to confirm linearity between absorbance and concentration. FT-IR spectroscopy is essential to assess EAC’s compatibility with hydrophilic polymers (PEG4000, PEO, Pectin, HA) by comparing spectra and ensuring safe, stable interactions with excipients (Figure-2-6).

Figure 2: FTIR spectrum of EAC

Figure 3: FTIR spectrum of EAC with PEG 4000

Figure 4: FTIR spectrum of EAC with PEO

Figure 5: FTIR spectrum of EAC with Pectin

Figure 6: FTIR spectrum of EAC with HA

Table 1: Formulation of EAC buccal films

Ingredients	F1	F2	F3	F4	F5	F6	F7	F8	F9
EAC Solid dispersion(mg)	16	16	16	16	16	16	16	16	16
PEO (mg)	200	300	400	-	-	-	-	-	-
Pectin (mg)	-	-	-	50	100	150	-	-	-
Hyaluronic acid (mg)	-	-	-	-	-	-	100	200	300
Glycerin (mL)	0.5	0.5	0.5	0.4	0.04	0.04	0.1	0.1	0.1
Starch (mg)	-	-	-	50	100	150	-	-	-
Tartaric acid (mg)	4	4	4	4	4	4	4	4	4
Poloxamer (mg)	2	2	2	2	2	2	2	2	2
Mannitol (mg)	4	4	4	4	4	4	4	4	4
Distilled water(mL)	10	10	10	10	10	10	10	10	10

EAC Solid Dispersion and Buccal Film Preparation:

EAC buccal films were prepared using solvent evaporation for solid dispersion⁸. A 1:2 ratios of EAC (500 mg) and PEG 4000 (1000 mg) was dissolved in methanol, stirred at 40˚C, and left to evaporate for 3 days. The dried mixture was ground, sieved (0.71 mm), and stored. For film casting, 5 mg of EAC solid dispersion was combined with polymers (PEO, Pectin, HA) and glycerine. The dispersion, mixed with additional excipients, was poured into petri dishes, air-dried for 24 hours, and oven-dried at 40°C. Films were then cut into 20 mm discs. (Table-1)

Evaluation of Buccal Films:^9-12

Buccal film quality control tests randomly choose 2 x 2 cm films. Physical qualities include visual inspection, average weight, weight fluctuation, thickness, folding endurance, swelling index, surface pH, drug content uniformity, and in-vitro drug release tests (Table-2). Morphological Assessment^13-16. Color, transparency, homogeneity, and surface qualities are examined on ten films (Figure-7).

Weight variation:

Ten films with different polymer concentrations are weighed separately and collectively on an electronic balance. The average weight is matched to film weights.

Thickness:

A screw gauge micrometer measures ten films from each formulation at four corners and center to calculate mean thickness and standard deviation.

Figure 7: Physical appearances of examples of different concentrations’ polymers (a) PEO (b) Pectin (c) HA formulated buccal films

Folding endurance:

Ten films are folded 180° till cracks emerge to measure flexibility. The mean and standard deviation are calculated from folds before cracking.

Measurement of Stickiness and peelabiltiy:

A buccal film is cut into uniform 2 x 2 cm pieces and put on pig buccal tissue or an artificial mucosal membrane to test its stickiness and peelability. A texture analyzer or tensile testing equipment measures the force needed to peel the film from the substrate at 1 mm/s. To determine the film's stickiness, the adhesive force (N/cm) and peel strength are measured.

Table 2: Weight variation, thickness, folding endurance, swelling index and surface pH of all formulations of buccal films

Code	Weight variation (g/cm²)	Thickness (mm)	Folding endurance	Swelling Index (%)	Surface pH
F1	0.0176 ± 0.0011	0.020 ± 0.007	755 ± 27	Dissolved in 3 min	6.68 ± 0.04
F2	0.0243 ± 0.0025	0.043 ± 0.009	825 ± 30		6.65 ± 0.05
F3	0.0310 ± 0.0029	0.048 ± 0.010	907 ± 31		6.67 ± 0.05
F4	0.0123 ± 0.0009	0.016 ± 0.007	510 ± 42	289.03 ± 48.95	6.73 ± 0.05
F5	0.0149 ± 0.0009	0.021 ± 0.006	602 ± 32	340.24 ± 46.26	6.71 ± 0.03
F6	0.0216 ± 0.0028	0.032 ± 0.006	777 ± 60	435.14 ± 90.28	6.68 ± 0.04
F7	0.0173 ± 0.0019	0.012 ± 0.004	602 ± 41	3.71 ± 0.35	6.72 ± 0.04
F8	0.0202 ± 0.0018	0.021 ± 0.007	500 ± 34	5.28 ± 0.20	6.70 ± 0.05
F9	0.0282 ± 0.0082	0.027 ± 0.007	423 ± 38	6.34 ± 0.73	6.67 ± 0.05

Swelling Index Studies:¹⁷

An important indicator of film moisture absorption is the swelling index. This study weighed 10 films as W1 and placed them in petri dishes with 5 mL of pH 6.8 isotonic phosphate buffer. The films were removed at 15, 30, 45, and 1hr intervals until a steady weight was attained. Swollen films were weighed as W2 after filter paper removed excess water. The swelling index was computed from W1 and W2.

Swelling Index (%) = W₂/(W1−W2) ×100%

Surface pH Assessment:¹⁸

The film's surface pH must be assessed to identify adverse effects since acidic or basic pH levels can irritate the mucosa. Swelling was allowed for 2 hours in petri dishes with 5 mL of pH 6.8 isotonic phosphate buffer solution and 10 selected films. After measuring the core surface pH of the swollen films with pH indicator sheets, the findings were analyzed using a standard color scale.

Drug Content Uniformity:^19-22

In this basket-method study using USP-II six-station paddle equipment, three films were attached to the central shaft with cyanoacrylate glue. Films were tested in pH 6.8 isotonic buffer at 37 ± 0.5˚C for 1 hour at 50 rpm. Three mL samples were collected hourly, replaced with fresh medium, filtered, diluted, and analysed via UV spectrophotometry at 421 nm (Table-3).

Table 3: Drug content uniformity of all formulations of buccal films (n = 3)

Formulation Code	% Drug Content
F1	99.05 ± 0.78
F2	93.09 ± 0.97
F3	87.39 ± 0.59
F4	90.23 ± 0.80
F5	89.07 ± 0.59
F6	81.81 ± 0.81
F7	95.42 ± 0.81
F8	99.05 ± 0.78
F9	93.09 ± 0.97

In vitro Drug Release study:

USP-compliant in vitro drug release studies for 2x2 cm buccal films were conducted in isotonic buffer (pH 6.8) at 37°C using a Franz diffusion cell or USP paddle device. Samples (2-5 mL) were periodically taken from the receptor compartment and replaced with fresh buffer at specified intervals. Drug concentration was determined using UV-Vis spectrophotometry at 421 nm, with cumulative drug release calculated and modeled (e.g., Hixson-Crowell) to analyse release kinetics. Triplicate studies were performed for reliability (Figure-8 and Table-4).

Figure 8: Hixson-Crowell Model of evaluation for Drug release studies

RESULTS AND DISCUSSIONS:

Compatibility Studies:

FTIR analysis assessed EAC-polymer compatibility, showing key peaks for EAC at 2884 cm⁻¹ (phenolic O-H), 1737 cm⁻¹ (C=O), and 1626 cm⁻¹ (C=C), which were consistent in EAC solid dispersion with PEG 4000 and buccal film polymers (PEO, Pectin, HA). Slight peak intensity changes for EAC + PEO and EAC + Pectin at 1738 cm⁻¹ and 2970 cm⁻¹ suggest minor C=O formation and phenolic O-H reduction. EAC and HA showed no intensity or peak shifts, indicating no significant chemical interactions with any studied polymer.

Buccal Film Quality Control Tests:

Buccal films with EAC solid dispersion, PEO, Pectin, HA, glycerine, tartaric acid, poloxamer, and mannitol passed stringent quality control testing to meet standards.

Visual or Physical Inspection:

Formulation physical properties were carefully monitored and recorded.

Surface Smoothness:

F1 to F3 had the smoothest surfaces, whereas F4 was smooth but rougher. Roughness was similar in F5–F9.

Homogeneity:

All formulations' homogeneity evaluations showed little orange particles in the films. EAC's low solubility and processing with a 0.71 mm sieve may cause this difficulty. Particle size reduction improves buccal film disintegration with a 0.63 mm sieve.

Peelability/stickiness:

F7 to F9 were sticky after detachment, whereas F1 to F6 were easily peelable. The HA polymer's characteristics may improve buccal mucosa adherence but make application difficult.

Table-4: In vitro drug release profiles of all formulations of buccal films. (n = 3)

Time (min)	Cumulative percentage drug release (%)
Time (min)	F1	F2	F3	F4	F5	F6	F7	F8	F9
0	0	0	0	0	0	0	0	0	0
1	19.8 ± 0.1	12.5 ± 0.2	6.7 ± 0.1	12 ± 0.2	13.1 ± 0.1	11.5 ± 0.1	18.2 ± 0.1	11.6 ± 0.2	7.9 ± 0.1
5	41.0 ± 0.2	4.0 ± 0.1	30.1 ± 0.3	20.3 ± 0.2	16.5 ± 0.4	16.1 ± 0.3	23.4 ± 0.2	17.2 ± 0.1	13.6 ± 0.3
8	48.3 ± 0.4	43.9 ± 0.5	44.4 ± 0.1	23.8 ± 0.2	24.7 ± 0.5	19.3 ± 0.3	42.6 ± 0.2	27.3 ± 0.2	22.3 ± 0.3
20	74.2 ± 0.6	71.9 ± 0.7	64.6 ± 0.4	49.6 ± 0.5	44.4 ± 0.3	41.1 ± 0.6	59.7 ± 0.4	54.1 ± 0.5	45.4 ± 0.2
45	87.4 ± 0.7	85.7 ± 0.5	76.8 ± 0.6	65.1 ± 0.6	64.5 ± 0.4	59.2 ± 0.5	83.1 ± 0.3	77.2 ± 0.6	71.6 ± 0.7
60	97.1 ± 0.1	94.6 ± 0.3	86.1 ± 0.3	86.2 ± 0.3	85.8 ± 0.4	80.6 ± 0.3	94.6 ± 0.28	89.7 ± 0.4	86.5 ± 0.2

Colour:

The color of EAC made all buccal films yellow (Figure 7).

Transparency:

The formulas were clear (Figure 7).

Physical-chemical properties:

Selected films were assessed for weight, thickness, flexibility, swellability, surface pH, drug content homogeneity, and in vitro drug release.

Weight Variation:²³

Table 4 shows minor standard deviation results for EAC loading weight fluctuations. Film weight increased with polymer content.

Thickness:²⁴

PEO, Pectin, and HA thickness measurements (Table 4) ranged from 0.020 to 0.048 mm, 0.016 to 0.032 mm, and 0.012 to 0.027 mm, with minor standard deviations. Film thickness increased with polymer content. Due to PEO's higher molecular weight, formulation F2 with PEO (300 mg) averaged 0.043 mm, topping F9 with HA (0.027 mm).

Flexibility:²⁵

Table 4 shows folding endurance as a measure of film flexibility: all films exceeded 300 folds without breaking, with PEO films at 755–907, Pectin at 510–777, and HA at 423–602. Higher PEO and Pectin concentrations improved flexibility, while increased HA concentration reduced durability. PEO's high plasticizer content contributed to its superior folding endurance.

Swelling Index Studies:

The swelling index, crucial for assessing mucoadhesion, indicated that PEO films (F1-F3) disintegrated within 3 minutes in isotonic phosphate buffer (pH 6.8), preventing index calculation. Pectin films (F4-F6) showed higher swelling indices (289.03 ± 48.95% to 435.14 ± 90.28%) compared to HA films (F7-F9), which exhibited much lower values (3.71 ± 0.35% to 6.34 ± 0.73%).

Polymer Concentration and Swelling Behaviour:

Increased polymer content in buccal films causes swelling, with PEO swelling and dissolving fastest due to its better water permeability (F1, F2, F3). Pectin films, especially F6 with the highest concentration of starch, swelled but may induce patient discomfort and delay EAC release due to prolonged buccal administration.

Surface pH and Compatibility Assessment:

Buccal films must be pH compatible with the mucosal membrane for maximum drug absorption and minimal discomfort. Films were incubated in pH 6.8 isotonic phosphate buffer for 2 hours to assess surface pH. Table 4 shows pH values between 6.6 and 6.7, match saliva's neutral pH (6.5 to 6.8). This shows less mucosal irritation, making the films more patient-friendly.

Drug Content Uniformity Assessment:

For medication content consistency, buccal films must distribute drugs uniformly. Drug concentration ranged from 81.81% to 99.05% across formulations (Table 3). PEO (F1-F3), Pectin (F4-F6), and HA (F7-F9) showed constant EAC content. Formulation F6 (81.81%) did not fulfill USP uniformity standards (85% to 115% of label claim), whereas the others varied from 85.57% to 99.05%.

In vitro Drug Release Analysis:²⁶

In vitro drug release tests were performed on three samples from each formulation using the USP-II dissolving paddle. As shown in Table 6 and Figure 10, formulation F1 had the maximum EAC release at 97.1±0.04% at 60 minutes. Polymer kinds and concentrations affected release.

Influence of Polymers on EAC Release Profiles²⁷:

Three polymers—PEO (F1, F2, F3), Pectin (F4, F5, F6), and HA (F7, F8, F9)—influence EAC release profiles, as shown in Figure 10. PEO formulations defeated Pectin and HA with cumulative EAC release percentages of 97.1%, 94.6%, and 86.1% in 60 minutes. HA released 33.3%, 27.9%, and 22%, whereas PEO released 48.3%, 43.9%, and 44.4% at 8 minutes. Pectin released 23.4%, 24.4%, and 19.8%. Due to polymers' water permeability and swelling, PEO released drugs faster than HA and pectin.

Influence of Plasticizer Concentration:

Glycerine concentration influences EAC release in buccal films, with 0.5 mL in PEO films creating voids for faster diffusion, enhancing release kinetics over Pectin (0.04 mL) and HA (0.1 mL). Higher polymer concentrations slow EAC release due to longer matrix wetting times, with high-swelling formulations F6 (Pectin) and F9 (HA) (Table 4). Drug release fits first-order kinetics, indicating concentration-dependent absorption. The Hixson-Crowell model (y = 0.056x, R² = 0.8866) suggests that dissolution surface area changes affect release.

CONCLUSION:

EAC's low hydrophilicity hinders buccal absorption, nevertheless this work shows that polymer type and concentration in film formulations can improve it. The best absorption improver is formulation F1, with 97.1% EAC release in 60 minutes. EAC release was sustained in Buccal films with Pectin and starch excipient. The study also stresses improving polymer type and concentration because higher polymer concentrations lower EAC release. PEO polymer released better than Pectin and HA at lower concentrations.

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